The specific aims of this proposal are to separate and isolate the multiple penicillin binding proteins of Escherichia coli and Staphylococcus aureus, to study their biochemical functions, the mechanism of their interactions with penicillins and cephalosporins and their relationship to inhibited bacterial enzymes (transpeptidases and D-alanine carboxypeptidases involved in bacterial cell wall synthesis) and to the penicillin killing site(s). Detailed chemical studies of their struture and of the mechanism of interaction of beta-lactam antibiotics with these substances will be undertaken including an effort to obtain the total structure of at least one penicillin bindng protein. These stuies represent the most detailed approaches possible to understanding the mecanism of interaction of a drug with its targets. There are no comparable studies with any other drug.